Mohiuddin SS and Manjrekar P. Interrelation Between Acute Phase Response with Copyright© Mohiuddin SS and Manjrekar P

Open Access Journal of Endocrinology ISSN: 2578-4641

Interrelation between Acute Phase Response with Altered Level of Serum Orosomucoid and Glycemic status in Newly Developed Adult as Well as Adolescent Diabetic Patients

Mohiuddin SS1* and Manjrekar P2 Research Article 1Department of Biochemistry, Imam Abdulrahman Bin Faisal University, Kingdom of Saudi Arabia Volume 2 Issue 3 Received Date: October 07, 2018 2Department of Biochemistry, Manipal University, India Published Date: October 19, 2018

*Corresponding author: Shamim Shaikh Mohiuddin, Assistant Professor, Department of Biochemistry, College of Medicine, Imam Abdulrahman Bin Faisal University, PO box- 1982, Dammam- 31441, Kingdom of Saudi Arabia, Email: [email protected]

Abstract

Orosomucoid is an acute phase protein and is associated with inflammation. Following an acute inflammatory episode increased levels of orosomucoid used to be observed. The clinical value of orosomucoid is currently limited to monitoring the acute phase reaction. The interrelation between cytokine mediated acute inflammatory response and elevation of plasma orosomucoid level of adult type 2 diabetic patients are almost fully established and well recognized but the correlation of inflammatory status with orosomucoid level with glycemic status in adolescent type 1 diabetic patients are still contradictory and under investigation. In this present study we tried to establish the interrelation of orosomucoid level as an indicator of acute inflammatory condition with glycemic status in case of adult as well as adolescent diabetic patients. The plasma levels of orosomucoid were determined for twenty-five newly diagnosed adult type 2 and twelve adolescent type 1 diabetic patients. Thirty normal controls were also chosen matching with age and sex with the patients. Although there are vast variation in results regarding the interrelation between the status of serum orosomucoid level as an low grade chronic inflammatory markers with glycemic status in case of adult as well as adolescent diabetic patient and but after the analysis of data obtained in our present study we can say with conviction that the interrelation of this level are well corelated in case of adult type 2 diabetic patient but in case of adolescent patients its still in contradictory phase and for definitive diagnosis it obviously required further study, investigation and follow up.

Keywords: Type 1 diabetes; Type 2 diabetics chronic low-grade inflammation; α1- acid glycoprotein; Orosomucoid

Interrelation between Acute Phase Response with Altered Level of Serum Orosomucoid and Glycemic status in Newly Developed Adult as Well as Adolescent Diabetic Patients J Endocrinol 2 Open Access Journal of Endocrinology

Introduction polypeptide moiety. It has an excellent solubility in water and polar solvents and has a high negative charge density Type 1 diabetes which is observing for about 90% of at pH 7.4. Treatment with neuraminidase followed by cases is the predominant form of diabetes during isoelectric focusing reveals polymorphism in the α1 childhood and adolescence, although increasing number region. Though polymorphism has no known clinical of obesity epidemic in childhood are often associated with significance but because of its orosomucoid migration on childhood onset type 2 diabetes [1,2]. The major agarose or cellulose electrophoresis may differ slightly etiological factors for onset of childhood type 1 diabetes from one specimen to another. Orosomucoid is associated mellitus are the synergistic effects of environmental, with inflammation. Following an acute inflammatory genetic and immunological factors which used to destruct episode increased levels of orosomucoid observed, i.e. it is the pancreatic β-cells mass. The main contributing factor an acute phase reactant. A certain degree of homology for occurrence of type 1 diabetes in individuals of genetic exists among the amino acid sequence of orosomucoid, susceptibility is due to process of autoimmunity that used the immunoglobins and haptoglobin α chain which to develop several months or even years which used to suggest a common ancestry or role of the immune system destroy the β-cells mass [3]. Complications of diabetes put [11]. Early work indicated that the liver was the only site a heavy burden for patients, family members and health of synthesis of orosomucoid. More recent studies have provider. The significant improvements in diabetes care shown that under certain well-defined condition, some used to achieve because of fighting against each individual tumors are able to synthesize orosomucoid. Catabolism of complication, especially for microvascular complications, orosomucoid proceeds first by desialation followed by yet macroangiopathy remains a major source of morbidity rapid degradation in liver within minutes. The clinical and mortality. By understanding of the complex value of orosomucoid determination is currently limited pathophysiology for development of adult as well as to monitoring the acute phase reaction. However childhood diabetes, prevention of the development of increased serum occurs in rheumatoid arthritis, systemic complication and treatment approach can be achieved [4]. lupus erythematosus, Crohn’s disease, and malignant Several inflammatory markers have been involved for neoplasms, especially those with metastasis and large detection of the incidence of diabetes, including C-reactive tumor masses and myocardial infarction. Decreased level protein (CRP) [5], sialic acid, white blood cells, interleukin occurs in malnutrition, severe hepatic damage and severe (IL) 6 [6] and orosomucoid [7] Several studies have protein loosing gastroenteropathy [11]. The acute-phase shown that circulating markers of inflammation, acute proteins are mainly synthesized in the liver, stimulated by phase reactants or interleukin-6 (IL-6) are strong cytokines, mainly interleukin (IL)-1, IL-6 and tumor predictors of the development of type 2 diabetes [8,9]. T-2 necrosis factor (TNF) which are produced in ((Type 2) diabetes mellitus is seen to be related with macrophages, monocytes, endothelium and many other increased blood concentrations of markers of the acute- cells in the body [12]. The acute-phase proteins have phase response known as α1 acid glycoprotein and also many activities that in general contribute to host defense, interleukin-6, which is the main cytokine mediator of the healing and adaptation to insult. For example, proteinase response. The dyslipidemia which is very much inhibitors such asa1-antitrypsin control proteinases re- commonly seen in Type 2 diabetes (hypertriglyceridemia leased by phagocytes, fibrinogen has a major role in and low serum levels of HDL cholesterol) is also a feature hemostasis, and ceruloplasmin, haptoglobin and of natural and experimental acute-phase reactions. Out of metallothionine are antioxidants protecting against toxic these all proteins our study of interest in this project is oxygen metabolites produced at the site of injury and orosomucoid, also known as α1-acid glycoprotein (AGP). inflammation [13].

Schmidt’s excellent review [10] covers historical Materials and Methods aspects of work that dates back the 1940’s when the seromucoid proteins were first described. The major Aims and Objective portion of plasma seromucoids has since been recognized 1. To detect the elevation of orosomucoid, if any, in newly as orosomucoid or α1- acid glycoprotein. Among the diagnosed untreated adult type 2 diabetic and plasma orosomucoid is unique because of its low pI which adolescent type 1 diabetes mellitus patient is 2.7-3.5. Its molecular weight is ~40000. It contains 45% 2. Compare the level of this inflammatory marker of carbohydrate as hexose, hexosamine and sialic acid in newly diagnosed untreated type 2 diabetes mellitus equal proportion. The antigenic determinant (epitope) with their glycemic status. utilized in immunochemical assays, however, is on the

Mohiuddin SS and Manjrekar P. Interrelation between Acute Phase Response with Copyright© Mohiuddin SS and Manjrekar P. Altered Level of Serum Orosomucoid and Glycemic status in Newly Developed Adult as Well as Adolescent Diabetic Patients. J Endocrinol 2018, 2(3): 000132.

3 Open Access Journal of Endocrinology

3. Compare the level of this inflammatory marker of using Folin Ciocalteau reagent. The last of these is newly diagnosed untreated type 1 diabetes mellitus employed in the technique given below. with their glycemic status. Statistics Participants The data was analyzed by the students’ t test and the Subjects were selected from various clinics and ANOVA test. Pearson’s coefficient was applied for hospitals in Mangalore, India. Height and weight of all correlational analysis. subjects were recorded, and body mass index was

calculated. None of the sixty-seven volunteers did not suffer from chronic inflammatory diseases like asthma, Results chronic bronchitis, and rheumatoid arthritis as was ascertained by clinical history. The study was approved The mean age (range), body mass index (BMI) and values of random blood sugar (RBS) are presented in by institutional ethical committee. Table 1. The control group participants were so chosen as

to cover the age range of the test groups. Table 2 lists the Materials values of orosomucoid in all three groups as mean ± SD. 5 ml blood was collected in plain bottle. Informed Table 3 denoted the comparison between different groups consent was taken from the individual subjects prior to and significance levels (p values). In newly diagnosed blood collection. Blood was taken from antecubital vein of type 2 diabetic patients (Group II) show higher level of the subjects and α-1 acid glycoprotein assay in serum was orosomucoid in compare to control group(Group III) as carried out by the method of Winzler RJ, et al. [14]. depicts the significant p value(< 0.0001*). It’s also detected that the level of the orosomucoid is significantly Principle of the Test higher in newly diagnosed type 2(Group II) in compare to newly diagnosed type 1(Group I) as denoted by After removing heat coagulable proteins with significant p value (< 0.0001*). Table 4 denoted perchloric acid, the orosomucoid which remains in the correlation of RBS (r value) with orosomucoid level in solution is precipitated by phosphotungstic acid and different groups of patients. This r value does not show estimated by determining it carbohydrate content by any positive correlation with any types of patients. reaction with an orcinol-sulphuric acid reagent, or its nitrogen by Kjeldahl nesslerization or its tyrosine content

Type 1 Diabetes Mellitus(N=12) Type 2 Diabetes Mellitus (N=25) Control(n=30)

(Mean ± SD) (Mean ± SD) (Mean ± SD) Age (years.) 18.33 ± 7.64 48.22 ± 7.11 44.97 ± 15.06 BMI 19.50 ± 1.23 24.03 ± 1.46 21.75 ± 2.27 RBS 338.25 ± 50.97 193.26 ± 35.30 94.20 ± 7.00 Table 1: The anthropometric data of the subjects participated in the study are presented in Table 1.

Type 1 Diabetes Type 2 Diabetes Mellitus(N=25) Control(N=30) Serum Level (Mg/Dl) Mellitus(N=12) (Mean ± SD) (Mean ± SD) (Mean ± SD) Orosomucoid(mg/dl) 94.87 ± 23.31 181.93 ± 31.94 103.41 ± 22.13 Table 2: The compare of mean value of orosomucoid in groups in Table 2.

Comparison between groups Level(mg/dl) Level(mg/dl) p value Comparison between Group I and Group III 94.87 ± 23.31(I) 103.41 ± 22.13(III) 0.275 Comparison between Group II and Group III 181.93 ± 31.94(II) 103.41 ± 22.13(III) < 0.0001* Comparison between Group I and Group II 94.87 ± 23.31(I) 181.93 ± 31.94(II) < 0.0001* Table 3: Comparison of level of orosomucoid (mg/dl) between different groups in Table-3 (p value < 0.05 is considered significant).

4 Open Access Journal of Endocrinology

Parameter Group I Group II Group III diabetic patients. These all previous work supported our present findings. Orosomucoid -0.15 -0.03 -0.05 In this present study if we analysis the value obtained Table 4: Correlation of RBS with orosomucoid in all the of serum orosomucoid in case of adult as well as groups (r value). adolescent patients it will shown that in case of untreated Group I = Type 1 diabetes mellitus patient (newly adult diabetic patients the values are much higher than diagnosed) adolescent diabetics. But if we concentrated only on Group II = Type 2 diabetes mellitus patient (newly random blood sugar level in both type of patients, it was diagnosed) determined much more higher level in case of adolescent Group III = Control patients (338.25 ± 50.97 mg/dl ) than in adult diabetic *denoted significant value patients(193.26 ± 35.30 mg/dl). In spite of this huge n = number of subjects difference of glycemic status, the level of inflammatory SD = Standard Deviation markers such as serum orosomucoid value is much higher BMI= Body Mass Index in case of adult untreated diabetic patients. From this RBS= Random Blood Sugar above finding we can predict that glycemic status of both untreated adolescent as well as adult diabetic used to not Discussion influenced by their current glycemic status. This finding is in accordance to early finding of Sriharan M, et al. [19]. In this present project we were aimed to determine Even it was also shown by Engstrom G, et al. [20], Schmidt whether serum orosomucoid levels are altered in case of MI, et al. [7], Duncan BB, et al. [21] and Pradhan AD, et al. adult as well as adolescent diabetic patients. The level of [22] that inflammatory markes used to elevated well serum orosomucoid were not elevated in twelve studied before the clinical manifestation of hyperglycemia cases of adolescent diabetic patients, in fact there were developed. This also supported the thought that glycemic significantly declined orosomucoid level (94.87 ± 23.31) status may not related to activated innate immune in these cases in compare to control (103.41 ± 22.13). system. But about decades before Allessandri G, et al. [23] Even p value also shown statistically insignificant (p shown that decrease plasma level of glucose decrease the 0.275). Some previous contradictory results were shown level of acute phase reactants. Even study done by by some renounced scientific works done by renounced Sriharan, et al. [19] shown positive correlation between scientists like Crooke MA, et al. [15], who was shown that inflammatory markers and 2 hours post prandial blood serum orosomucoid levels used to decline in case of glucose levels. The main basic mechanism for the adolescent diabetic patients which is supporting our augmented response needs to be well understand and findings. But contradictory result was shown by Gomes, et stimulus of the response should be finding it out. Several al. [16]. In their study there were significantly increase in numbers of hypothesis put forwards in support of this serum value of orosomucoid in case of adolescent diabetic mechanism by Pickup JC, el at. [24] and Grimble RF, et al. patients. [25], which included resistance to insulin, obesity, atherosclerosis, other complications of diabetes and In case of twenty-five newly diagnosed adult diabetic maladaptation of normal innate immune mechanism due patient there were significant elevation of serum to environmental threats. It was also shown by orosomucoid level observed. Even p value was Mohammad AV, et al. [26] that in post prandial state determined < 0.0001 in compare with controls, which adipocytes secret number of proinflammatory cytokines. used to consider as statistically significant. This result had Later it was proposed by ‘common soil theory’ which used a co- agreement with most of the authors worked to evaluate the association of inflammatory markers with previously on orosomucoid level on adult diabetic diabetes and development of atherosclerosis. patients and shown inflammatory markers tends to Hyperglycemia and insulin resistance could promote increase in adult diabetic patients. McMillan DE [17] inflammation and inflammation may be a factor linking shown that serum orosomucoid level used to increase in diabetes mellitus to the development of atherosclerosis. long termed adult diabetic patients as well as Festa A, et Bayens JW and Thorpe SR [27] and even Brownlee M [28] al. [18] shown that elevated levels of acute phase proteins shown that elevated level of serum glucose promotes and plasminogen activator inhibitor 1 predict the inflammatory response by promoting oxidative stress by development of in case of insulin resistance type 2 increasing the level of tumor necrosis factor kappa B. In this study present the mean BMI was found to be 19.5 ±

5 Open Access Journal of Endocrinology

1.23 in adolescent patient and 24.03 ± 1.46 in adult 6. Pradhan AD, Manson JE, Rifai N, Buring JE, Ridker PM patients. No correlation was found between BMI and (2001) Creactive protein, interleukin 6, and risk of serum orosomucoid levels. developing type 2 diabetes mellitus. JAMA 286(3): 327-334. Conclusion 7. Schmidt MI, Duncan BB, Sharrett AR, Lindberg G, After analysis of whole study hence we can come to an Savage PJ, et al. (1999) Markers of inflammation and initial conclusion that multiple factors are involved in prediction of diabetes mellitus in adults development of innate immune response. From these (Atherosclerosis Risk in Communities study): a above findings we can say in conviction that the role of cohort study. Lancet 353(9165): 1649-1652. low grade chronic inflammatory markers such as orosomucoid in case of adult diabetic patients are almost 8. Snijder MB (2001) C-reactive protein and diabetes beyond doubt but its role in case of adolescent diabetic type 2. Diabetologia 44(1): 115. patients are still in contradictory phase, although the 9. Spranger J, Kroke A, Mohlig M, Hoffmann K, course of progression of disease and resulting Bergmann MM, et al. (2003) Inflammatory cytokines complications in both type of diseases is almost same. Out and the risk to develop type 2 diabetes: results of the of all one of a most fatal complication is developing of prospective population based European Prospective atherosclerosis resulting in cardiovascular disease. So we Investigation Cancer and Nutrition (EPICN)- Potsdam can come to a conclusion that although level of study: Diabetes 52(3): 812-818. inflammatory markers such as orosomucoid is well associated and correlated with adult diabetes mellitus but 10. Schmid K (1975) α-1 acid glycoprotein. In: FW much further work and follow up can be done to establish Putnam (Ed.), The plasma proteins. 2nd(Edn.), New the definitive relation of level of inflammatory markers York, Academic Press, pp: 184-228. such as orosomucoid in pathogenesis of adolescent diabetic patients. 11. Tietz NW (1986) Amino acids and proteins. In: Textbook of clinical chemistry. WB Saunders References Company, pp: 519-618.

1. Craig ME, Hattersley A, Donaghue KC (2009) 12. Kushner I (1993) Regulation of the acute phase Definition, epidemiology and classification of diabetes response by cytokines. Perspect Biol Med 36(4): 611- in children and adolescents. Pediatr Diabetes 10(12): 622. 3-12. 13. Taniuchi K, Chifu K, Hayashi N, Nakamachi Y, 2. Patterson CC, Dahlquist GG, Gyurus E, Green A, Soltesz Yamaguchi N, et al. (1981) A new enzymatic method G, et al. (2009) Incidence trends for childhood type 1 for the determination of sialic acid and its application diabetes in Europe during1989-2003 and predicted as a marker of acute phase reactants. Kobe J Med Sci new cases 2005-20: a multicentre prospective 27(3): 91-102. registration study. Lancet 373(9680): 2027-2033. 14. Winzler RJ (1995) Determination of serum α-1 acid 3. Pugliese A (1995) Unraveling the genetics of insulin glycoprotein. In: Methods in Biochemical Analysis. dependent type I diabetes: The search must go on. Interscience Pub New York 2: 270. Diabetes Rev 7(1): 39-54. 15. Crook MA, Tutt P, Simpson H, Pickup JC (1993) Serum 4. Sartore S, Agostini C, Avogaro A (2007) Significance sialic acid and acute phase protein in type 1 and type of endothelial progenitor cells in subjects with 2 diabetes. Clin Chim Acta 219(1-2): 131-138. diabetes. Diabetes Care 30(5): 1305-1313. 16. Gomes MB, Piccirillo LJ, Nogueira VG, Matos HJ (2003) 5. Thorand B, Lowel H, Schneider A, Kolb H, Meisinger C, Acute phase proteins among patients with type1 et al. (2003) C-reactive protein as a predictor for diabetes. Diabetes Metab 29(4 pt 1): 405-411. incident diabetes mellitus among middle-aged men: results from the MONICA Augsburg cohort study, 17. McMillan DE (1989) Increased levels of acute phase 1984-1998. Arch Intern Med 163(1): 93-99. serum proteins in diabetes. Metabolism 38(11): 1042-1046.

6 Open Access Journal of Endocrinology

18. Festa A, D Agostino R, Tracy RP, Haffner SM (2002) of developing type2 diabetes mellitus. JAMA 286(3): Elevated levels of acute phase proteins and 327-334. plasminogen activator inhibitor 1 predict the development of type 2 diabetes: the insulin resistance 23. Allessandri G, Raju K, Gullino PM (1983) Mobilization atherosclerosis study. Diabetes 51(4): 1131-1137. of capillary endothelium in vitro induced by effectors of angiogenesis in vivo. Cancer Res 43(4): 1790-1797. 19. Sriharan M, Reichelt AJ, Opperman ML, Duncan BB, Mengue SS, et al. (2002) Total sialic acid and 24. Pickup JC, Crooke MA (1998) Is type 2 diabetes associated elements of the metabolic syndrome in mellitus a disease of the innate immune system? women with and without previous gestational Diabetologia 41(10): 1241-1248. diabetes. Diabetes Care 25(8): 1331-1335. 25. Grimble RF (2002) Inflammatory status and insulin 20. Engstrom G, Stavenow L, Hedblad B, Lind P, Eriksson resistance. Curr Opin Cli Nutr Metab 5(5): 551-559. KF, et al. (2003) Inflammation sensitive plasma protein, diabetes mortality and incidence of 26. Mohammed AV, Goodrick S, Rawesh A, Katz DR, Miles myocardial infarction and stroke: A population based JM, et al. (1997) Subcutaneous adipose tissue releases study. Diabetes 52(2): 442-447. interleukin-6, but not tumor necrosis factor α, in vivo. J Clin Endocrinol Metab 82(12): 4196-4200. 21. Duncan BB, Schmidt MI, Offenbacher S, Wu KK, Savage PJ, et al. (1999) Factor VIII and other 27. Bayens JW, Thorpe SR (1999) Role of oxidative stress hemostasis variables are related to incident diabetes in diabetic complications: a new perspective on an old in adult: The Atherosclerosis Risk In Community paradigm. Diabetes 48(1): 1-9. (ARIC) study. Diabetes Care 22(5): 767-772. 28. Brownlee M (2001) Biochemistry and molecular cell 22. Pradhan AD, Manson JE, Rifai N, Buring JE, Ridker PM, biology of diabetic complication. Nature 414 (6865): et al. (2001) C-reactive protein, interleukin-6 and risk 813-820.